COMPLEXES BETWEEN CHAPERONIN GROEL AND THE CAPSID PROTEIN OF BACTERIOPHAGE-HK97

The 42 kDa capsid protein of bacteriophage HK97 requires the GroEL and GroES chaperonin proteins of its Escherichia coil host to facilitate correct folding, both in vivo and in vitro. In the absence of GroES an d ATP, denatured gp5 forms a stable complex with the 14 subunit GroEL molecule. We characterized the electrophoretic and biochemical propert ies of this complex. In electrophoresis on a native (nondenaturing) ge l, the band of the gp5-GroEL complex shifts to a slower migrating posi tion relative to uncomplexed GroEL. The results show that there is onl y one subunit of gp5 bound to each GroEL 14-mer and that the shift in band position is due primarily to a change in the overall charge of th e complex relative to uncomplexed GroEL, and not to a change in size o r shape. GroEL forms similar complexes with proteolytic fragments of g p5, with a series of sequence duplication derivatives of gp5, and with other proteins. Electrophoretic examination of these complexes shows that a band shift occurs with proteins larger than 31-33 kDa but not w ith smaller proteins. For those proteins that cause a band shift upon complex formation, the magnitude of the shift is correlated with the p redicted negative charge on the protein; paradoxically, the direction of the band shift is opposite to what is predicted if the charge of th e complex were simply the sum of the charge of GroEL and the charge of the substrate protein. We suggest that binding of a substrate protein to GroEL is accompanied by a net binding of solution cations to the c omplex, but only in the case of proteins above a minimum size of 31-33 kDa. The gp5-GroEL complex is in an association/dissoication equilibr ium, with a binding constant measured in the range of 11-17 mu M(-1).